Despite significant process in ubiquitin modification by using traditional genetic methods, chemical small molecules that directly target and modify ubiquitin are little reported. Here, we find that a fluorescigenic pyrazoline derivative (FPD5) could do so effectively. Molecule docking revealed that lysine 11 of ubiquitin was the key contact residue. FPD5, with stronger fluorescence, elevated the ubiquitination of beclin 1 (BECN1) and promoted autophagy. This study highlights that targeting ubiquitin by chemical small molecules enables us to modulate ubiquitination and the downstream signaling in the ubiquitin system. (C) 2020 Elsevier Inc. All rights reserved.